Rates of denitrification (isotope pairing) and nitrogen fixation (acetylene reduction) were simultaneously measured in three temperate, intertidal Zostera muelleri meadows and adjacent non-vegetated tidal flats within Western Port, Australia. Net daily nitrogen fluxes ranged from -276 (net denitrification) to 520 μmol N/m2 d (net nitrogen fixation), and were generally positive, with only two instances of net negative fluxes. The highest fluxes were observed at the sites with the lowest water column nitrate concentrations. No significant differences in net nitrogen fluxes were found between vegetated and non-vegetated sediments (p=0.213). Nitrogen fixation was generally the dominant process occurring, which was stimulated in the presence of vegetation except at the most marine-influenced site, where nitrogen fixation in non-vegetated sediments was higher. Nitrogen fixation rates in non-vegetated sediments were highly correlated to cyanobacterial cell counts (although no mats were present). Rates were ∼65 μmol N/m2 d at 0 cell counts, suggesting a basal rate driven within the sediment. Additional slurry experiments confirmed significant rates of nitrogen fixation within the sediment, which were stimulated by sucrose and terminated by nitrate (p<0.05), strongly suggesting sulfate-reducing bacteria contributed to nitrogen fixation. At the bay-wide scale, nitrogen fixation was estimated to contribute ∼430 t N/yr compared to ∼650 t N/yr from catchment and atmospheric inputs and 230 t N/yr lost through denitrification. Sensitivity analysis confirmed that while the loss of seagrass would affect the magnitude of the bay-wide flux of nitrogen on these tidal flats, nitrogen fixation remains the dominant process.